Direct gene transfer into rat articular cartilage by in vivo electroporation

Grossin, Laurent; Cournil, Christel; Mir, Lluis M.; Liagre, Bertrand; Dumas, Dominique; Étienne, S.; Guingamp, Corinne; Netter, Patrick; Gillet, Pierre

doi:10.1096/fj.02-0518com

Cited by 43 publications

(39 citation statements)

References 28 publications

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“…Thus, one can imagine a biological targeting of inflammation sites via the homing at inflammatory sites of lymphocytes pre-transfected in the spleen. In order to reach local production of cytokines too, a study with the GFP reporter gene demonstrated the feasibility of electroporation to the knee after intraarticular injection [90]. Until now, most of the studies realized on other organs, such as cornea [14], tendon [12], liver [11], bladder [13], brain [17], etc., have focused on vector development and establishment of transfer techniques.…”

Section: Electrotransfer In Other Tissuesmentioning

confidence: 99%

Plasmid DNA electrotransfer for intracellular and secreted proteins expression: new methodological developments and applications

et al. 2004

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SummaryIn vivo electrotransfer is a physical method of gene delivery in various tissues and organs, relying on the injection of a plasmid DNA followed by electric pulse delivery. The importance of the association between cell permeabilization and DNA electrophoresis for electrotransfer efficiency has been highlighted. In vivo electrotransfer is of special interest since it is the most efficient non-viral strategy of gene delivery and also because of its low cost, easiness of realization and safety. The potentiality of this technique can be further improved by optimizing plasmid biodistribution in the targeted organ, plasmid structure, and the design of the encoded protein. In particular, we found that plasmids of smaller size were electrotransferred more efficiently than large plasmids. It is also of importance to study and understand kinetic expression of the transgene, which can be very variable, depending on many factors including cellular localization of the protein, physiological activity and regulation. The most widely targeted tissue is skeletal muscle, because this strategy is not only promising for the treatment of muscle disorders, but also for the systemic secretion of therapeutic proteins. Vaccination and oncology gene therapy are also major fields of application of electrotransfer, whereas application to other organs such as liver, brain and cornea are expanding. Many published studies have shown that plasmid electrotransfer can lead to long-lasting therapeutic effects in various pathologies such as cancer, blood disorders, rheumatoid arthritis or muscle ischemia. DNA electrotransfer is also a powerful laboratory tool to study gene function in a given tissue.

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Section: Electrotransfer In Other Tissuesmentioning

confidence: 99%

Plasmid DNA electrotransfer for intracellular and secreted proteins expression: new methodological developments and applications

et al. 2004

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“…Gene therapeutics lead to an inherent sustained release of expressed therapeutic genes by host cells in situ, which is more effective due to higher bioactivity of the hostproduced growth factor (Bonadio et al, 1999;Bleiziffer et al, 2007). Plasmid DNA has been successfully applied in direct in vivo gene transfer for wound healing and angiogenesis (Michlits et al, 2007;Mittermayr et al, 2008), cardiac regeneration (Sundararaman et al, 2011) and musculoskeletal regeneration (Grossin et al, 2003;Kawai et al, 2006;Osawa et al, 2009;Osawa et al, 2010). Taking these advantages into account, non-viral vectors, such as plasmid DNA, are considered the most likely candidates for clinical translation of tissue regenerative gene therapies.…”

Section: Introductionmentioning

confidence: 99%

Constitutive and inducible co-expression systems for non-viral osteoinductive gene therapy

Feichtinger

Hacobian²,

Hofmann³

et al. 2014

eCM

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Tissue regenerative gene therapy requires expression strategies that deliver therapeutic effective amounts of transgenes. As physiological expression patterns are more complex than high-level expression of a singular therapeutic gene, we aimed at constitutive or inducible co-expression of 2 transgenes simultaneously.Co-expression of human bone morphogenetic protein 2 and 7 (BMP2/7) from constitutively expressing and doxycycline inducible plasmids was evaluated in vitro in C2C12 cells with osteocalcin reporter gene assays and standard assays for osteogenic differentiation. The constitutive systems were additionally tested in an in vivo pilot for ectopic bone formation after repeated naked DNA injection to murine muscle tissue.Inductor controlled differentiation was demonstrated in vitro for inducible co-expression. Both co-expression systems, inducible and constitutive, achieved significantly better osteogenic differentiation than single factor expression. The potency of the constitutive co-expression systems was dependent on relative expression cassette topology. In vivo, ectopic bone formation was demonstrated in 6/13 animals (46 % bone formation efficacy) at days 14 and 28 in hind limb muscles as proven by in vivo µCT and histological evaluation.In vitro findings demonstrated that the devised single vector BMP2/7 co-expression strategy mediates superior osteoinduction, can be applied in an inductor controlled fashion and that its efficiency is dependent on expression cassette topology. In vivo results indicate that co-expression of BMP2/7 applied by non-viral naked DNA gene transfer effectively mediates bone formation without the application of biomaterials, cells or recombinant growth factors, offering a promising alternative to current treatment strategies with potential for clinical translation in the future.

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“…The majority of these studies were gene therapy studies, such as siRNA or protein delivery into chondrogenic cell lines, or, animal models of arthritis [11][12][13] . In other systems, such as chicken or mouse, electroporation of facial mesenchyme has been challenging (personal communications, Dept of Craniofacial Development, KCL).…”

Section: Introductionmentioning

confidence: 99%

Electroporation of Craniofacial Mesenchyme

Tabler

Liu

2011

JoVE

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Electroporation is an efficient method of delivering DNA and other charged macromolecules into tissues at precise time points and in precise locations. For example, electroporation has been used with great success to study neural and retinal development in Xenopus, chicken and mouse [1][2][3][4][5][6][7][8][9][10] . However, it is important to note that in all of these studies, investigators were not targeting soft tissues. Because we are interested in craniofacial development, we adapted a method to target facial mesenchyme.When we searched the literature, we found, to our surprise, very few reports of successful gene transfer into cartilaginous tissue. The majority of these studies were gene therapy studies, such as siRNA or protein delivery into chondrogenic cell lines, or, animal models of arthritis [11][12][13] . In other systems, such as chicken or mouse, electroporation of facial mesenchyme has been challenging (personal communications, Dept of Craniofacial Development, KCL). We hypothesized that electroporation into procartilaginous and cartilaginous tissues in Xenopus might work better. In our studies, we show that gene transfer into the facial cartilages occurs efficiently at early stages (28), when the facial primordium is still comprised of soft tissue prior to cartilage differentiation.Xenopus is a very accessible vertebrate system for analysis of craniofacial development. Craniofacial structures are more readily visible in Xenopus than in any other vertebrate model, primarily because Xenopus embryos are fertilized externally, allowing analyses of the earliest stages, and facilitating live imaging at single cell resolution, as well as reuse of the mothers 14 . Among vertebrate models developing externally, Xenopus is more useful for craniofacial analysis than zebrafish, as Xenopus larvae are larger and easier to dissect, and the developing facial region is more accessible to imaging than the equivalent region in fish. In addition, Xenopus is evolutionarily closer to humans than zebrafish (˜100 million years closer) 15 . Finally, at these stages, Xenopus tadpoles are transparent, and concurrent expression of fluorescent proteins or molecules will allow easy visualization of the developing cartilages. We anticipate that this approach will allow us to rapidly and efficiently test candidate molecules in an in vivo model system.

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Direct gene transfer into rat articular cartilage by in vivo electroporation

Cited by 43 publications

References 28 publications

Plasmid DNA electrotransfer for intracellular and secreted proteins expression: new methodological developments and applications

Plasmid DNA electrotransfer for intracellular and secreted proteins expression: new methodological developments and applications

Constitutive and inducible co-expression systems for non-viral osteoinductive gene therapy

Electroporation of Craniofacial Mesenchyme

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